The functional role of exosomal miRNA-675-3p derived from patients with hepatocellular carcinoma.

Abstract

Background: Hepatocellular carcinoma (HCC) is one of the most common solid tumors, resulting in poor survival and high mortality worldwide. MicroRNAs (miRNAs) contained in serum exosomes are being increasing used as targets for disease diagnosis and treatment. However, the role and the diagnostic potential of exosome-transported miRNAs in HCC remain largely underexplored. Therefore, this study aims to identify differentially expressed exosomal miRNAs in the serum of HCC patients and healthy individuals, clarify the biological function of the key miRNAs in HCC progression, and explore its potential as a diagnostic and therapeutic biomarker for HCC.

Methods: In this study, the serum-derived exosomes of patients with HCC and healthy individuals were characterized via transmission electron microscopy (TEM), Western blot assay, and nanoparticle tracking analysis (NTA). The serum-derived exosomes were labeled with PKH-67, and the transport to recipient HCC cells was observed with confocal microscopy. Differentially expressed miRNAs (DEmiRs) encapsulated in serum-derived exosomes were screened via miRNA sequencing, the expression of which was verified through quantitative real-time polymerase chain reaction (qRT-PCR). The association between miR-675-3p and clinicopathological features was investigated via the chi-squared test. The gain and loss of function of miR-675-3p in HCC cells were examined through transfection with mimics and inhibitor. Cell Counting Kit-8 (CCK8), colony formation, 5-ethynyl-2'-deoxyuridine (EdU) staining, and Transwell assays were conducted to test the biological effects of overexpressed and inhibited miR-675-3p on HCC cells in vitro. Tumor xenograft models and immunohistochemistry (IHC) were performed to assess the malignant progression of HCC cells with downregulated miR-675-3p in vivo.

Results: Uptake of exosomes from patients with HCC promoted the malignant phenotype of recipient cells. miRNA-675-3p within exosomes was confirmed to be upregulated in the serum of patients with HCC as compared with that of healthy donors. The chi-squared test results showed that miRNA-675-3p was associated with poor survival. Overexpression of miRNA-675-3p promoted the proliferation, invasion, and colony formation of HCC cells in vitro, while downregulation of miRNA-675-3p had the opposite biological effects on HCC cells and inhibited tumor growth and malignant progression in vivo. Furthermore, bioinformatics analysis indicated that miRNA-675-3p was involved in certain cellular processes, especially in lipid metabolism and regulation of several signaling pathways such as PI3K/AKT pathways and PPAR pathways.

Conclusions: miRNA-675-3p was upregulated in the serum of patients with HCC and exerted an oncogenic role in HCC cells via the targeting of downstream genes. These findings may constitute novel insights into the development of HCC, with miRNA-675-3p potentially serving as a biomarker for the diagnosis and treatment of patients with HCC.